Radioactively labelled amino acid analogues, their preparation and use

ABSTRACT

The present invention relates to Halogenated amino acid analogues for use in diagnosis, which compounds have the genera formula (I) wherein: R is (C 1 -C 6 ) alkyl optionally substituted with thioether or ether oxygen atom when n=0, or a substituted aromatic or heteraromatic ring when n=1-6; and m=0 or 1; and X is a halogen atom. The invention further relates to precursor compounds for these analogues, to a method of preparing these analogues, to a pharmaceutical composition comprising these analogues and to the use of these analogues and compositions in the diagnosis of cancer.

FIELD OF THE INVENTION

The present invention relates to amino acid analogues labelled withhalogen atom, such as a radioactive fluorine atom, such as F-18, or anon-radioactive fluorine atom, such as F-19. The invention furtherrelates to precursor compounds for and a method of preparing theseanalogues, to a pharmaceutical composition comprising these analoguesand to the use of this composition for diagnosis, for example by meansof Positron Emission Tomography or functional MRI.

BACKGROUND OF THE INVENTION

Whatever the new approaches for therapy of cancers will be in thefuture, an accurate and specific non-invasive diagnosis on bio-molecularlevel of tumours and metastases will remain of primary importance.Transformation of normal cells into malignant cells is caused by changesin the genetic material, resulting in subtle but fundamental metabolicchanges like increased glucose metabolism and increased amino aciduptake and metabolism. These changes in the metabolic phenotype permitthe in-vivo study of tumours using radioactive labelled tracers coupledto SPECT (Single Photon Emission Computed Tomography) or PET (PositronEmission Tomography). PET linked coincidence acquisition allows a betterresolution and quantification than SPECT, needed for tumour tracing anddimensioning.

Currently, the use of ¹⁸F-FDG (fluoro-deoxyglucose) and PET is the mostimportant technique in nuclear medicine for the study of oncologypatients. Although this method is very sensitive, it has two majorlimitations, namely an avid accumulation in inflammatory lesions andhigh uptake in the brain, jeopardizing the diagnosis of brain tumours.

It was shown that the use of radioactive amino acids for SPECT and PETcould overcome these shortcomings for the larger part. In the late 80's,several ¹¹C-labelled amino acids like methionine and tyrosine, as wellas 2-¹⁸F-tyrosine (2-¹⁸F-Tyr) of high specific activity were used forPET studies. At that time it was believed that a high specific activitywas required and that for tumour-specification the labelled amino acidhad to be involved in a high rate protein incorporation. None of theseamino acids has meanwhile been introduced into routine clinical PETbecause of the short half life and insufficient in vivo stability ofC-11 or complicated radiochemical synthesis resulting in insufficientyield (such as for 2-¹⁸F-Tyr).

About the same time, L-3-¹²³I-alpha-methyl-tyrosine (3-¹²³I-IMT) wasintroduced as a SPECT tracer for brain tumours and is used until nowalso for other tumours like sarcoma and lymphoma. The uptake of thistracer in tumours occurs for the larger part by the L transport system.The plasma membrane transport system L is in many cells the only(efficient) pathway for the import of large branched and aromaticneutral amino acids. The L-type amino acid transporter 1 (LAT1) is a Na⁺independent amino acid transporter and is over-expressed inover-expressed in malignant cell as it plays a critical role in cellgrowth and proliferation. For functional expression LAT1 requires theheavy chain of the surface antigen 4F2 (heavy chain 4F2hc). Theincreased accumulation is mainly determined by strongly increased aminoacid transport activity rather than incorporation into proteins. A majordrawback limiting the applicability of this tracer is however the highrenal accumulation.

O-(2-¹⁸F-ethyl)-tyrosine (FET) and ¹⁸F-alpha-methyl-tyrosine wereproposed in 1999 as potential PET tracers. The compounds showed the sameuptake properties as IMT. The preparation of these tracers stillrequires complicated and time consuming synthetic steps and HPLC stepslimiting the overall radiochemical yield. They are therefore in practicenot very useful.

In the research that led to the invention two new potential SPECTtracers, 2-¹²³I-tyrosine (2-¹²³I-Tyr) and 2-¹²³-I-phenylalanine, weredeveloped. When evaluated in vivo in RIM tumour(rhabdomyo-sarcoma)-bearing rats, these tracers showed high uptake inthe tumours (comparable with IMT) while no renal accumulation (10 timesless activity in the kidneys than IMT) or high brain uptake wasobserved. Kinetic studies also revealed that the uptake of radioactiveamino acid reflected the amounts of amino acids in the tumour ascompared to the blood pool compartment and that no high specificactivity is required for the tracer. However, also these tracers arealmost limited to SPECT as the positron emitting iodine isotopes ¹²⁴Iand ¹²²I do not have the required radionuclide properties for routinepatient PET diagnosis.

SUMMARY OF THE INVENTION

It was found that a ¹⁸F-labelled amino acid as tumour tracer showshigher tumour specificity as compared to FDG and is better suited asbrain tracer. The fact that within toxicity limits neither high specificactivity nor non-carrier added preparation of the ¹⁸F-tracer isrequired, should allow for electrophilic radio-fluorination making useof [¹⁸F]—F₂. However, the radioisotope production yield with thecurrently available F₂-targets is limited and even with an almostquantitative labelling yield, amounts comparable with those of the¹⁸F-FDG production are far from being reached and does not allow routinemulti patient PET diagnosis.

It is therefore the object of the present invention to provide newcompounds and precursors therefor that can be easily and quicklysynthesized and can thus also be labelled with F-18 which has ahalf-life of only 2 hours. It is a further object of the invention toprovide the use of such compounds in diagnosis.

The inventors considered based on the results obtained with ¹⁸F-FET andtheir own results with 2-¹²³I-Phe and 2-¹²³I-Tyr that the aromatic aminoacid properties are conserved after substitution of an O-ethyl group andeven in the presence of a voluminous iodine atom. This invention is thusbased on the new approach to introduce an alkyl side chain on the phenylring to facilitate introduction of the radioactive atom. They thusprovided an ¹⁸F-alkyl-phenyl structure in phenylalanine and tyrosine,either ortho, meta or para. Examples are ¹⁸F—CH₂-Phe or ¹⁸F—CH₂—CH₂-Pheand 2-¹⁸F—CH₂-Tyr or 2-¹⁸F—CH₂—CH₂-Tyr. This reduces the labellingchemistry to direct conventional nucleophilic aliphatic substitution onthe alkylphenylic side branch of the L-amino acid. In this approachcumbersome stereospecific synthesis is not required. The same strategywas followed for the radio-fluorination of the aliphatic amino acidsleucine and isoleucine. Preliminary uptake experiments in R1M cells invitro in a buffer simulating in vivo conditions, showed for ³H-leucineand ³H-isoleucine results comparable with 3H-Tyr and ³H-Phe. Sincealiphatic-substituted F hardly changes the pharmacology, it follows thatthese aliphatic amino acids are also suitable molecules forradio-fluorination.

DETAILED DESCRIPTION OF THE INVENTION

The invention thus relates to halogenated amino acid analogues havingthe he general formula

wherein:R is (C₁-C₆)alkyl optionally substituted with thioether or ether oxygenatom when n=0, or a substituted aromatic or heteraromatic ring whenn=1-6; and m=0 or 1; andX is a halogen atom.

R is preferably an alkyl selected from methyl, ethyl, propyl, isopropyl,butyl, tertiary butyl or methyl thioethyl ether when n is 0 and R ispreferably phenyl, hydroxyphenyl, pyridyl, hydroxypyridinyl when n is 1,2 or 3.

The radioactive halogen atom is preferably a radioactive fluorine, inparticular ¹⁸F because of its radionuclidic properties which makes itwithin the positron emitting isotopes the most interesting for labellingtracer molecules for diagnosis with PET.

Suitable amino acid analogues of the invention are analogues of thearomatic or heteroaromatic amino acids phenylalanine, tyrosine andazatyrosine or the alkyl amino acids alanine, valine, leucine,isoleucine and methionine. The aromatic amino acids are preferablyderivatized at the 2 position (phenyl) and 3 position (2-pyridylanalogue) with a (C₁-C₂)alkyl methyl and ethyl. The alkyl can also bepresent at the 3 and 4 position on the aromatic ring of phenylalanineand 5 position in meta-tyrosine.

Preferred analogues are selected from the group consisting of [¹⁸F]labelled β-2-fluoromethylphenyl-α-aminopropionic acid, [¹⁸F] labelledβ-3-fluoromethylphenyl-α-aminopropionic acid, [¹⁸F] labelledβ-4-fluoromethylphenyl-α-aminopropionic acid, [¹⁸F] labelledβ-2-fluoroethylphenyl-α-aminopropionic acid, [¹⁸F] labelledβ-3-fluoroethylphenyl-α-aminopropionic acid, [¹⁸F] labelledβ-4-fluoroethylphenyl-α-aminopropionic acid, [¹⁸F] labelledβ-2-fluoromethylphenyl-α-aminopropionic acid, [¹⁸F] labelledβ-3-fluoromethyl-2-pyridyl-α-aminopropionic acid, [¹⁸F] labelledβ-4-fluoromethyl-2-pyridyl-α-aminopropionic acid, [¹⁸F] labelledβ-5-fluoromethyl-2-pyridyl-α-aminopropionic acid, [¹⁸F] labelledβ-3-fluoroethyl-2-pyridyl-α-aminopropionic acid, [¹⁸F] labelledβ-4-fluoroethyl-2-pyridyl-α-aminopropionic acid, [¹⁸F] labelledβ-4-fluoroethyl-2-pyridyl-α-aminopropionic acid, [¹⁸F] labelled2-amino-3-(5-fluoromethyl-3-hydroxyphenyl)propianoic acid, [¹⁸F]labelled 2-amino-3-(6-fluoromethyl-3-hydroxyphenyl)propianoic acid,[¹⁸F] labelled 2-amino-3-(2-fluoromethyl-4-hydroxyphenyl)propianoicacid, [¹⁸F] labelled2-amino-3-(2-fluoroethyl-5-hydroxypyridyl)propianoic acid, [¹⁸F]labelled 2-amino-3-(3-fluoroethyl-5-hydroxy-2-pyridyl)propianoic acid,[¹⁸F] labelled 2-amino-3-(5-fluoroethyl-3-hydroxyphenyl)propianoic acid,[¹⁸F] labelled alanine, [¹⁸F] labelled valine, [¹⁸F] labelled leucine,[¹⁸F] labelled isoleucine and [¹⁸F] labelled methionine. Of these theanalogues of which the 2 OR 6 position of the aromatic ring issubstituted with the alkyl are found to be preferred because the 4position (para) is not sterically hampered for biochemical recognition.The invention also relates to all of the above compounds that carry anon-radioactive label, in particular a non-radioactive fluorine atom.

The invention further relates to a pharmaceutical composition comprisingone or more amino acid analogues as claimed and an excipient, carrier ordiluent. The excipient, diluent or carrier can be any compound orcomposition in liquid form, that is sterile and non-pyrogenic and can beisotonic saline or an isotonic buffer.

The pharmaceutical composition can be used as a tracer in PositronEmission Tomography (PET) and functional MRI.

The invention further relates to the use of the amino acid analogues inthe preparation of a pharmaceutical composition for the diagnosis ofcancer.

According to another aspect thereof the invention provides a method fordiagnosing a patient for the presence of tumours and/or metastases,which comprises administration of a diagnostic effective amount of oneor more of the amino acid analogues, and visualising the localisation ofthe analogues in the patients body, such as by means of PET, orfunctional MRI.

The present invention further provides precursor compounds for preparingthe amino acid analogues, which precursors have the general formula

wherein:R is (C₁-C₄) alkyl when n=0 or phenyl or pyridyl when n=1, 2 or 3;X is a leaving group, in particular tosyl, mesityl triflate or ahalogen; andNH₂ and COOH are protected.

The substitution of an alkyl group, provided with an appropriate leavinggroup, on the phenyl ring of an aromatic amino acid; such asphenylalanine or tyrosine, or introduction of a leaving group on thealiphatic side chain of alkyl amino acid allows for introduction of theradioactive atom, in particular fluorine, such as ¹⁸F, by aliphaticnucleophilic substitution. This is a quick synthesis step allowing ahigh radioactive labelling yield.

The COOH may be esterified with a (C₁-C₅)alkyl. The (C₁-C₅)alkyl isselected from the group consisting of methyl, ethyl, propyl, isopropyl,tertiary butyl, neopentyl. NH₂ may be protected with a group selectedfrom N-Boc, N-trityl, f-moc or others. The technology of protecting withthese compounds is well known to the person skilled in the art and forexample described in Protecting Groups in Organic Synthesis, T. W.Greene, John Wiley & Sons, 1981.

In the precursor compounds R is preferably methyl, ethyl, propyl,isopropyl, isobutyl, 1-methyl butyl, methyl thioethyl ether when n is 0and R is preferably phenyl, hydroxyphenyl, pyridyl, hydroxypyridyl whenn is 1, 2 or 3.

The halogen that may be used as a leaving group in the precursormolecules may be a “cold”, i.e non-radioactive halogen.

Suitable precursor compounds of the invention are analogues of thearomatic amino acids phenylalanine and tyrosine, the hetero aromaticazatyrosine or the alkyl amino acids alanine, valine, leucine,isoleucine and methionine. The aromatic amino acids are preferablyderivatized at the 2 position (phenyl) or the 3 position (pyridyl) withan (C₁-C₂) alkyl, such as methyl, ethyl, so that the 4 position (para)is not sterically hindered for biochemical recognition. The alkyl canalso be present at the 5 position on the aromatic ring in meta-tyrosine.

The precursor molecules based on alkylated aromatic amino acids can beprepared starting from commercially available alkyl amino acids such asL-2-CH₃-Phe. L-2-CH3-Phe will be protected by esterification (^(t)But)and as N-Boc or N-trityl and radicalar mono-bromination or iodination ofthe 2-methyl group is performed. A tosyl (Tos), mesityl (Mes) or atriflate (Trif) group and any other suitable leaving group is introducedby nucleophilic exchange. After purification, the compound is storedunder nitrogen.

As L-/D-2-Br-Phe is commercially available, precursor compounds with Bras the halogen can also be obtained by a Wurtz-Fittig reaction, usingdibromomethane and then applying the same pathways as described above.

L-2-Tos(Trif)-CH₂-Tyr can be prepared starting from CH₃O-L-2-I-Tyr,which is commercially available and is an adequate precursor for theWurtz-Fittig pathway mentioned above.

For the synthesis of L-/D-2-(Tos, Mes, Trif)ethyl-Phe, L-/D-4-(Tos, Mes,Trif)ethyl-Phe, L-/D-2-(Tos, Mes, Trif) methyl-Tyr and L-/D-2-(Tos, Mes,Trif) ethyl-Tyr, the same strategies are followed.

For Val, Leu and Ile a place specific bromination is applied, followedby introduction of the appropriate leaving group.

The invention further relates to a method for preparing the amino acidanalogues of the invention comprising substitution of the leaving groupwith a radioactive halogen atom. The substitution may take place bymeans of aliphatic nucleophilic substitution of tosyl, mesityl ortriflate with a radioactive halogen, in particular fluorine, or by meansof exchange of the halogen leaving group with a radioactive halogen, inparticular a radioactive fluoride.

When the aliphatic nucleophilic substitution of tosyl, mesityl ortriflate or non-isotopic exchange is used for preparing theradioactively labelled amino acid analogues this will result in acarrier-free preparation, because after substitution the radioactivemolecules are separated from the precursors. In case the isotopicexchange method is used a carrier-added preparation is obtained. Thespecific activity of this preparation depends on the amount ofnon-radioactive precursor present.

The amino acid analogues and precursor compounds of the invention canhave the L and D orientation. The method of the invention for producingthe labelled amino acid analogues can use either L or D compounds ormixtures thereof as the starting material thus leading to either L or Danalogues or mixtures thereof.

The present invention will be further illustrated in the Examples thatfollow and that are not intended to limit the invention in any way.Reference is made to the following figures:

FIG. 1: Inhibition of ³H-Phe/Phe uptake in R1M cells in vitro byL-2-F-methyl-PHE. The common intercept proves that the inhibition iscompetitive and that L-2-F-methyl-Phe uses the same transport system asL-Phe.

FIG. 2: PET of R1M tumour bearing rat. The tumour is visible at theupper right, the pancreas in the middle and the bladder under. 120 MbqL-2-¹⁸F-methyl-phenylalanine was intravenously injected.

EXAMPLES Example 1

Synthesis of Precursor Molecules and Non-Radioactive FluorinatedAnalogues.

1.1. Protected L-2-bromomethyl-PHE

On L-2-methyl-Phe the tributyl ester and N-Boc protection is introducedby conventional chemistry (N-Boc: (BOC)₂O, TEA, MeOH/tButOH, roomtemperature, 2 hours; Butylester: TMSL+tButOH or Li—O-t-butyl, roomtemperature, 24 hours). The protected compound is reacted in CCl₄ withBr-succinimide in the presence of benzoylperoxide as catalyst (radicalhalogenation) at 80 EC during 1 hour. After precipitation of thesuuccinimide the product is purified by column chromatography.

As alternative for benzoylperoxide an irradiated polymer like PMMA isused as radical promotor, this allows the purification by simplefiltration.

1.2. Protected L-2-Tosethyl-Phe

L-2-I-Phe is obtained by Cu1+ assisted iodo for bromo exchange oncommercial available L-2-Br-Phe in acidic reducing aqueous condition(gentisic acid and SnSO₄ as reducing agent for CuSO₄). Protection isintroduced as in 1.1. The ethyltosyl is introduced in 3 steps (a:vinylbromide, Pd(PPh₃)₄, 1,4-dioxane, 100 EC, 1 hour; b: BH₃-THFcomplex, 4N NaOH, 30% H₂O₂, THF, 0 EC, 2 hours; c: TsCl, DMAP, CH₂Cl₂,room temperature, 2 hours).

1.3. Protected Brominated Leucine

Protection is performed as described in 1.1. and 1.2. Radicalbromination is performed as described in 1.1.

1.4. Protected L-2-alkyltosyl-Tyrosine

L-2-I-Tyr is commercially available. The chemistry is the same asdescribed for L-2-I-phenylalanine in 1.2.

1.5. Protected Bromoleucine

N-Boc, t-Butyl protected leucine is brominated by a radical reaction asdescribed in 1.1.

1.6. Non Radioactive Fluorinated Analogues

These are obtained by reaction at reflux temperature of the Tosylatedprecursor molecules with nBu₄NF in CH₃CN.

Example 2

Radiochemical Synthesis of Compounds of the Invention

L-/D-¹⁸F—R-Phe analogues (R=methyl or ethyl) are prepared bynucleophilic exchange of ¹⁸F on L-/D-2-TosR-Phe in an AcN/TBA/HCO₃ ⁻ orAcN/K₂₂₂/CO₃ ²⁻ mixture at 85 EC during 5 minutes.

In short, ¹⁸F- is separated from the target water via an anion exchangecolumn. Elution of the activity is achieved with tetra-n-butyl ammoniumhydrogenc arbonate in H₂O. H₂O is discarded by azeotropic distillationafter addition of acetonitrile. L-2-Tosethyl-N-trityl-phenylalaninetert. butylester in dry acetonitrile is added to the ¹⁸F-recipient andheated during 3-5 minutes at 85 EC. After the reaction the solvent isevaporated by means of pre-heated N₂.

Then, two pathways are possible. First, de-esterification andde-protection are preformed in solution followed by HPLC or mini-columnpurification. Alternatively, straightforward de-protection can beperformed on a mini-column followed by HPLC or another type mini-columnpurification.

For L-/D-¹⁸F-Leu and L-/D-¹⁸F-Ile an analogous radiochemistry isapplied.

Example 3

In Vitro Affinity for Cancer Cells

The affinity of L-2-F-methyl-phenylalanine for uptake by the L-transportsystem 1 (LAT1) in cancer cells (rat rhabdomyo-sarcoma cells) wasdetermined by measuring the inhibition of the uptake ofL-³H-phenylalanine after 15 minutes incubation in HEPES buffer of pH 7.4containing appropriate amounts of L-phenylalanine and ofL-2-F-methyl-phenylalanine. The uptake was saturable and followed thetypical Michaelis-Menten relation allowing to draw Lineweaver-Burk(FIG. 1) plots.

The double reciprocal plots in FIG. 1 with a common intercept almost onthe 1/uptake axis shows that the inhibition is competitive with thephenylalanine uptake and uses the same LAT transporter system.

A mean Ki value of 76 :M was obtained for L-2-F-methyl-phenylalanine.This value is almost comparable with the Km value of 65 :M obtained forthe natural L-phenylalanine in the same conditions.

Example 4

In Vivo Rat Evaluation of L-2-18F-methyl-phenylalanine in a R1M TumourBearing Rat by Means of PET

FIG. 2 shows that high uptake is observed in the tumour and pancreas.The latter is typical for rodent. It shows that the¹⁸F-methyl-phenylalanine analogue is transported as a natural aminoacid.

No accumulation in the kidneys or other organs is observed. The productis cleared through the kidneys to the bladder.

1. A halogenated amino acid analogue having the general formula:

wherein: X is a radioactive halogen; m is 0 or 1, n is 0, 1, 2, 3, 4, 5,or 6; R is (C₁-C₆)alkyl optionally substituted with thioether or etheroxygen atom when n is 0; and R is a substituted aromatic orheteroaromatic ring when, n is 1, 2, 3, 4, 5 or
 6. 2. The analogue ofclaim 1, wherein R is methyl, ethyl, propyl, isopropyl, butyl, tertiarybutyl or methyl thioethyl.
 3. The analogue of claim 1, wherein R isphenyl, hydroxyphenyl, pyridyl or hydroxypyridyl.
 4. (canceled)
 5. Theanalogue of claim 4, wherein the halogen is ¹⁸F.
 6. The analogue ofclaim 4, wherein the halogen is ¹²³I. 7-8. (canceled)
 9. The analogue ofclaim 1, wherein the analogue is selected from the group consisting of:[¹⁸F] labeled L,D-2-amino-3-(2-fluoromethyl-phenyl)-propionic acid;[¹⁸F] labeled L,D-2-amino-3-(3-fluoromethyl-phenyl)-propionic acid; [¹⁸Flabeled L,D-2-amino-3-(4-fluoromethyl-phenyl)-propionic acid; [¹⁸F]labeled L,D-2-amino-3-(2-fluoroethyl-phenyl) propionic acid; [¹⁸F]labeled L,D-2-amino-3-(3-fluoroethyl-phenyl)propionic acid; [¹⁸F]labeled L,D-2-amino-3-(4-fluoroethyl-phenyl)-propionic acid; [¹⁸F]labeled L,D-2-amino-3-(3-fluoromethyl-pyridin-2-yl)propionic acid; [¹⁸F]labeled L,D-2-amino-3-(4-fluoromethyl-pyridin-2-yl)-propionic acid;[¹⁸F] labeled L,D-2-amino-3-(5-fluoromethyl-pyridin-2-yl)-propionicacid; [¹⁸F] labeledL,D-2-amino-3-(6-fluoromethyl-pyridin-2-yl)-propionic acid; [¹⁸F]labeled L,D-2-amino-3-(3-fluoroethyl-pyridin-2-yl)-propionic acid; [¹⁸F]labeled L,D-2-amino-3-(4-fluoroethyl-pyridin-2-yl)-propionic acid; (¹⁸F]labeled L,D-2-amino-3-(5-fluoroethyl-pyridin-2-yl)-propionic acid; [¹⁸F]labeled L,D-2-amino-3-(6-fluoroethyl-pyridin-2-yl)-propionic acid; [¹⁸F]labeled L,D-2-amino-3-(2-fluoromethyl-4-hydroxy-phenyl)-propionic acid;[¹⁸F] labeled L,D-2-amino-3-(5-fluoromethyl-3-hydroxy-phenyl)-propionicacid; [¹⁸F] labeledL,D-2-amino-3-(6-fluoromethyl-3-hydroxy-phenyl)-propionic acid; [¹⁸F]labeled L,D-2-amino-3-(2-fluoroethyl-4-hydroxy-phenyl)-propionic acid;[¹⁸F] labeled L,D-2-amino-3-(5-fluoroethyl-3-hydroxy-phenyl)-propionicacid; [¹⁸F] labeledL,D-2-amino-3-(6-fluoroethyl-3-hydroxy-phenyl)-propionic acid; [¹⁸F]labeled L,D-2-amino-3-(3-fluoromethyl-5-hydroxy-pyridin-2-yl)-propionicacid; [¹⁸F] labeledL,D-2-amino-3-(3-fluoroethyl-5-hydroxy-pyridin-2-yl)-propionic acid;[¹⁸F] labeledL,D-2-amino-3-(3-fluoromethyl-6-hydroxy-pyridin-2-yl)-propionic acid;[¹⁸F] labeledL,D-2-amino-3-(4-fluoromethyl-6-hydroxy-pyridin-2-yl)-propionic acid;[¹⁸F] labeledL,D-2-amino-3-(3-fluoroethyl-6-hydroxy-pyridin-2-yl)-propionic acid;[¹⁸F] labeledL,D-2-amino-3-(4-fluoroethyl-6-hydroxy-pyridin-2-yl)-propionic acid;[¹⁸F] labelled alanine; [¹⁸F] labelled valine; [¹⁸F] labelled leucine;[¹⁸F] labelled isoleucine; and [¹⁸F] labelled methionine.
 10. Apharmaceutical composition comprising the analogue of claim 1 and atleast one of an excipient, carrier and diluent.
 11. The pharmaceuticalcomposition of claim 10, wherein the pharmaceutical composition is usedas a tracer in at least one of Positron Emission, Tomography (PET) andfunctional Magnetic Resonance Imaging (MRI). 12-24. (canceled)